Factors Affecting the Efficiency of In Vitro Embryo Production in Prepubertal Mediterranean Water Buffalo

SIMPLE SUMMARY: Embryos can be produced from prepubertal animals using laparoscopic ovum pickup combined with in vitro embryo production technologies. However, due to their young age and unique reproductive physiology, there is currently very limited knowledge about what factors may affect the success of these programs in Mediterranean water buffalo. Here, we assessed how results are affected by season, age, individual variation and choice of sire used for fertilization. Specifically, we found that season and age only had limited impacts, but there were large variations between individual donors and sires. In addition, we assessed what factors can affect pregnancy rates after transferring the resulting embryos into recipient animals and found that regardless of number of embryos transferred, fresh or vitrified, all variables assessed were compatible with the establishment of pregnancies. ABSTRACT: Embryos from prepubertal water buffalo can be produced using laparoscopic ovum pickup (LOPU) and in vitro embryo production (IVEP). However, to date, it is unclear what factors and environmental conditions can affect LOPU-IVEP efficiency in prepubertal animals, especially buffalo. In this study, we explored the impact of season, age and individual variation among female donor animals, as well as the effect of the sire used for in vitro fertilization. Donor animals between 2 and 6 months of age were stimulated using gonadotropins prior to LOPU, which was performed at two-week intervals. Following in vitro maturation and fertilization, the resulting embryos were then cultured to the blastocyst stage until they were either vitrified or transferred into recipient animals. The number of follicles available for aspiration and embryo development rates was stable throughout the year. As animals became older, there was a slight trend for fewer COCs recovered from LOPU and better embryo development. There was a large individual variation in both ovarian response and the developmental competence of oocytes among donors. The bull used for fertilization also had a significant impact on embryo development. Upon embryo transfer, pregnancy rates were not affected by the number of embryos transferred per recipient. The best pregnancy rates were achieved when transferring blastocysts, compared to compact morula or hatched blastocysts. Finally, vitrification had no effect on pregnancy rate compared to fresh embryos.